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| 2. |
- Chen, X., et al.
(författare)
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Inhibition of Wnt/beta-catenin signaling suppresses bleomycin-induced pulmonary fibrosis by attenuating the expression of TGF-beta 1 and FGF-2
- 2016
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Ingår i: Experimental and Molecular Pathology. - : Elsevier BV. - 0014-4800. ; 101:1, s. 22-30
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Tidskriftsartikel (refereegranskat)abstract
- Pulmonary fibrosis is a progressive lung disorder of unknown etiology, which is characterized by alterations in alveolar epithelium function, fibroblast activation, and increased extracellular matrix deposition. Recent studies have demonstrated that PF is associated with uncontrolled production of cytokines after lung injury. In the present study, we found that transforming growth factor-beta 1 (TGF-beta 1) and fibroblast growth factor 2 (FGF-2) were both upregulated in bleomycin-induced fibrotic lung tissue and primary murine alveolar epithelial Type II (ATII) cells treated with bleomycin. Furthermore, we discovered that TGF-beta 1 could induce the differentiation of lung resident mesenchymal stem cells (LR-MSCs) into fibroblasts, which may play an essential role in PF. LR-MSCs incubated with FGF-2 showed modest alterations in the expression of alpha-SMA and Vimentin. Moreover, in our study, we found that Wnt/beta-catenin signaling was activated both in vitro and in vivo as a result of bleomycin treatment. Interestingly, we also found that suppression of the Wnt/beta-catenin signaling could significantly attenuate bleomycin-induced PF accompanied with decreased expression of TGF-beta 1 and FGF-2 in vitro and in vivo. These results support that controlling the aberrant expression of TGF-beta 1 and FGF-2 via inhibition of Wnt/beta-catenin signaling could serve as a potential therapeutic strategy for PF. (C) 2016 Elsevier Inc. All rights reserved.
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| 3. |
- Cheng, Wei Kang, et al.
(författare)
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Nuclear and stromal expression of Manic fringe in renal cell carcinoma
- 2021
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Ingår i: Experimental and molecular pathology (Print). - : Elsevier. - 0014-4800 .- 1096-0945. ; 122
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Tidskriftsartikel (refereegranskat)abstract
- Renal cell carcinoma (RCC) is the most common type of kidney cancer and has the highest mortality rate among genitourinary cancers. Despite the advances in molecular targeted therapies to treat RCC, the inevitable emergence of resistance has delineated the need to uncover biomarkers to prospectively identify patient response to treatment and more accurately predict patient prognosis. Fringe is a fucose specific beta 1, 3N-acetylglucosaminyltransferase that modifies the Notch receptors. Given the link between its function and aberrant Notch activation in RCC, Fringe may be implicated in this disease. The Fringe homologs comprise of Lunatic fringe (LFng), Manic fringe (MFng) and Radical fringe (RFng). MFng has been reported to play a role in cancer. MFng is also essential in the development of B cells. However, the expression profile and clinical significance of MFng, and its association with B cells in RCC are unknown. CD20 is a clinically employed biomarker for B cells. This pilot study aimed to determine if MFng protein expression can be utilized as a prospective biomarker for therapeutics and prognosis in RCC, as well as to determine its association with CD20+ B cells. Analysis of publicly available MFng gene expression datasets on The Cancer Genome Atlas Netlwork (TCGA) identified MFng gene expression to be up-regulated in Kidney Clear Cell Renal Carcinoma (KIRC) patients. However there was no significant association between the patient survival probability and the level of MFng expression in this cohort. Immunohistochemistry performed on a tissue microarray containing cores from 64 patients revealed an elevated MFng protein expression in the epithelial and stromal tissues of RCC compared to the normal kidney, suggesting a possible role in tumorigenesis. Our study describes for the first time to our knowledge, the protein expression of MFng in the nuclear compartment of normal kidney and RCC, implicating a prospective involvement in gene transcription. At the cellular level, cytoplasmic MFng was also abundant in the normal kidney and RCC. However, MFng protein expression in the malignant epithelial and stromal tissue of RCC had no positive correlation with the patients' overall survival, progression-free survival and time to metastasis, as well as the gender, age, tumor stage and RCC subtype, indicating that MFng may not be an appropriate prognostic marker. The association between CD20+ B cells and epithelial MFng was found to approach borderline insignificance. Nonetheless, these preliminary findings may provide valuable information on the suitability of MFng as a potential therapeutic molecular marker for RCC, thus warrants further investigation using a larger cohort.
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| 6. |
- Granqvist, Victoria, et al.
(författare)
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The combination of TRAIL and the Smac mimetic LCL-161 induces an irreversible phenotypic change of MCF-7 breast cancer cells
- 2022
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Ingår i: Experimental and Molecular Pathology. - : Elsevier BV. - 0014-4800. ; 125
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Tidskriftsartikel (refereegranskat)abstract
- Introduction: Breast cancer is the most common malignancy affecting women. Although the prognosis generally is good, a substantial number of patients still suffer from relapse, emphasizing the need for novel treatments. Smac mimetics were developed to facilitate cell death by blocking inhibitor of apoptosis proteins (IAPs). It has been suggested that TNF-related apoptosis inducing ligand (TRAIL) can be used together with Smac mimetics to induce cancer cell death. Methods: Cell viability was studied with Trypan blue staining and Annexin V assay, siRNA was used to downregulate specific proteins, protein levels were estimated with Western blot, and mRNA levels were analyzed with qPCR, microarray and RNA-seq. For global expression, groups were compared with principal component analysis and the limma package in R. Gene enrichment was analyzed with Fisher's test. For other experiments, significance of difference was tested by one-way ANOVA, followed by Tukey's HSD test. Results: The combination of Smac mimetic LCL-161 and TRAIL induces an irreversible change in phenotype, but not cell death, of luminal MCF-7 breast cancer cells. The cells become small and circular and dissociate from each other and the effect could not be reversed by returning the cells to regular growth medium. The morphology change could be prevented by caspase inhibition using z-VAD-FMK and downregulation of caspase-8. Caspase-7 is also indicated to be of importance since downregulation of this caspase resulted in fewer morphologically changed cells. Enrichment analyses of changes in global gene expression demonstrated that genes associated with estrogen receptor (ER) signaling are downregulated, whereas nuclear factor kappa B- (NF-κB) and interferon- (IFN) driven genes are upregulated in altered cells. However, inhibition of these pathways did not influence the change in morphology. Induction of IFN-induced genes were potentiated but NF-ĸB-driven genes were slightly suppressed by caspase inhibition. Conclusions: The results demonstrate that LCL-161 and TRAIL can irreversibly alter the MCF-7 breast cancer cell phenotype. However, the changes in morphology and global gene expression are mediated via separate pathways.
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| 11. |
- Nilsson, Harriet, et al.
(författare)
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CFTR and tight junctions in cultured bronchial epithelial cells
- 2010
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Ingår i: Experimental and molecular pathology (Print). - : Elsevier BV. - 0014-4800 .- 1096-0945. ; 88:1, s. 118-127
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Tidskriftsartikel (refereegranskat)abstract
- Airway epithelial salt and water transport takes place through paracellular and transcellular pathways. This transport depends critically on the epithelial sodium channel (ENaC) and the cystic fibrosis transmembrane conductance regulator (CFTR), operating in concert with the paracellular pathway through the tight junctions (TJs). Normal (16HBE14o-), cystic fibrosis (CFBE41o-), and corrected CFBE41o- (CFBE41o pCep4) airway epithelial cell lines were cultured under isotonic conditions. Transepithelial electrical resistance (TEER) was measured as indicator of the tightness of the cultures. Morphology was investigated by immunofluorescence and paracellular permeability by lanthanum nitrate or [14C] mannitol as permeability markers. CFBE41o pCep4 cells developed lower TEER than CFBE41o- cells. Addition of a specific inhibitor of CFTR (CFTRinh-172) to 16HBE14o- and CFBE pCep4 cells resulted in a time-dependent increase in TEER whereas stimulation of CFTR by IBMX and forskolin caused a decrease. Permeability to lanthanum and [14C] mannitol was lower in 16HBE14o- cells exposed to CFTRinh-172 and in CFBE41o- cells compared to untreated 16HBE14o- and CFBE41o pCep4 cells, respectively. 16HBE14o- cells exposed to IBMX and forskolin showed higher permeability to lanthanum but lower permeability to [14C] mannitol compared to control. Immunofluorescence revealed a disorganisation of F-actin and a-tubulin in 16HBE14o- cells exposed to CFTRinh-172, which was not seen in untreated cultures. A higher degree of disorganised F-actin and a-tubulin was also seen in CFBE41o- cells compared to CFBE41o- pCep4 cells. Changes in F-actin and a-tubulin in 16HBE14o- cells exposed to IBMX and forskolin were also seen, although these were less apparent. These results suggest the possibility of an interaction between the activity of CFTR and the TJ protein complex, probably via the cytoskeleton.
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| 12. |
- Oliynyk, Igor, 1974-, et al.
(författare)
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The effect of NO-donors on chloride efflux, intracellular Ca2+ concentration and mRNA expression of CFTR and ENaC in cystic fibrosis airway epithelial cells
- 2013
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Ingår i: Experimental and molecular pathology (Print). - Maryland Heights, USA : Elsevier. - 0014-4800 .- 1096-0945. ; 94:3, s. 474-480
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Tidskriftsartikel (refereegranskat)abstract
- Since previous studies showed that the endogenous bronchodilator, S nitrosglutathione (GSNO), caused amarked increase in CFTR-mediated chloride (Cl−) efflux and improved the trafficking of CFTR to the plasmamembrane, and that also the nitric oxide (NO)-donor GEA3162 had a similar, but smaller, effect on Cl− efflux, itwas investigatedwhether the NO-donor properties of GSNOwere relevant for its effect on Cl− efflux fromairwayepithelial cells. Hence, the effect of a number of other NO-donors, sodium nitroprusside (SNP), S-nitroso-Nacetyl-DL-penicillamine (SNAP), diethylenetriamine/nitric oxide adduct (DETA-NO), and diethylenetriamine/nitricoxide adduct (DEA-NONOate) on Cl− efflux from CFBE (ΔF508/ΔF508-CFTR) airway epithelial cells was tested.Cl− efflux was determined using the fluorescent N-(ethoxycarbonylmethyl)-6-methoxyquinoliniu bromide(MQAE)-technique. Possible changes in the intracellular Ca2+ concentration were tested by the fluorescent fluo-4method in a confocal microscope system. Like previously with GSNO, after 4 h incubation with the NO-donor, anincreased Cl− efflux was found (in the order SNAP > DETA-NO > SNP). The effect of DEA-NONOate on Cl− effluxwas not significant, and the compound may have (unspecific) deleterious effects on the cells. Again, as withGSNO, after a short (5 min) incubation, SNP had no significant effect on Cl− efflux. None of the NO-donors thathad a significant effect on Cl− efflux caused significant changes in the intracellular Ca2+ concentration. After 4 hpreincubation, SNP caused a significant increase in the mRNA expression of CFTR. SNAP and DEA-NONOatedecreased the mRNA expression of all ENaC subunits significantly. DETA-NO caused a significant decrease only inα-ENaC expression. After a short preincubation, none of the NO-donors had a significant effect, neither on theexpression of CFTR, nor on that of the ENaC subunits in the presence and absence of L-cysteine. It can be concludedthat the effect of GSNO on Cl−efflux is, at least in part, due to its properties as an NO-donor, and the effect is likely tobe mediated by CFTR, not by Ca2+-activated Cl− channels.
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| 13. |
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| 14. |
- Servetnyk, Zhanna, et al.
(författare)
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Chloride transport in NCL-SG3 sweat gland cells : channels involved
- 2007
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Ingår i: Experimental and molecular pathology (Print). - : Elsevier BV. - 0014-4800 .- 1096-0945. ; 83:1, s. 47-53
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Tidskriftsartikel (refereegranskat)abstract
- The aim of the study was to assess whether NCL-SG3, the only immortalized sweat gland cell line available, can be used as an in vitro model to study chloride ion transport in cultured sweat gland cells. Cl− efflux was measured using the MQAE dye fluorescence technique after stimulating the cells with different agonists. A significant stimulation of chloride efflux was achieved with the calcium ionophore A23187 resulting in an efflux rate of 0.9 mM/s. Both ATP and UTP activated chloride efflux in these cells, with the ATP response being larger. IBMX and forskolin stimulation did not induce a rate of chloride efflux above the basal level. Immunocytochemistry showed no detectable CFTR in NCL-SG3 cells. This finding was confirmed with flow cytometry analysis. Niflumic acid (20 and 100 μM NFA) and 4,4′-diisothiocyanatodihydrostilbene-2,2′-disulfonic acid (H2DIDS) (100 ìM) decreased the rate of ATP-stimulated chloride efflux significantly (0.40 and 0.31 mM/s with NFA, 0.37 mM/s with H2DIDS). Gadolinium (20 ìM) had no effect on the chloride transport rate. In conclusion, the NCL-SG3 cells retain some of the aspects of human sweat gland epithelium, such as the ability to form cell–cell contacts. The CFTR protein is neither functional nor expressed in cultured NCL-SG3 sweat gland cells. Ca2+-activated chloride conductance is confirmed and the putative Ca2+-activated chloride channel (CaCC) is further characterized in term of its pharmacological sensitivity. The NCL-SG3 sweat gland cell line can be used to investigate the characteristics of the CaCC and to identify the channel.
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| 15. |
- Servetnyk, Zhanna, et al.
(författare)
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The effect of S-nitrosoglutathione and L-cysteine on chloride efflux from cystic fibrosis airway epithelial cells
- 2011
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Ingår i: Experimental and molecular pathology (Print). - : Elsevier BV. - 0014-4800 .- 1096-0945. ; 90:1, s. 79-83
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Tidskriftsartikel (refereegranskat)abstract
- The endogenous bronchodilator, S-nitrosoglutathione (GSNO), has been proposed as a possible pharmacological remedy that reverses the Delta F508-CFTR (cystic fibrosis transmembrane conductance regulator) maturation defect and increases CFTR-mediated chloride efflux in cultured cystic fibrosis airway epithelial cells (CFBE41o(-)). It has also been reported that L-cysteine enhanced S-nitrosothiol uptake and increased the intracellular S-nitrosothiol levels, likely through transnitrosation chemistry. The present study investigated whether L-cysteine augmented the effect of GSNO on chloride efflux from CF airway epithelial cells. Treatment with 10 mu M GSNO combined with 20 mu M L-cysteine resulted in increased chloride efflux from CFBE41o(-) cells after 5 minutes exposure compared to the control efflux rate and to the efflux rate in the presence of L-cysteine alone as measured using the fluorescent dye N-(ethoxycarbonylmethyl)-6-methoxyquinolinium bromide (MQAE). Chloride efflux rates from these cells after 4 h exposure to GSNO and L-cysteine were not different from control. Treatment with 10 mu M GSNO alone increased chloride efflux from CFBE41o(-) cells after 4 h but not at shorter incubation times. GSNO with or without L-cysteine did not alter epithelial tight junction integrity. In conclusion, a combination of GSNO with L-cysteine led to significant increase in chloride efflux in CFBE41o(-) cells but the effect was transient and not sustained beyond minutes.
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